As a group, viruses have played major roles in both the number and significance of newly recognized diseases in both animals and humans. A recent example is Australian Bat Lyssavirus (ABLV). ABLV is a newly recognized rabies virus-related rhabdovirus belonging to the genus Lyssavirus and is an enveloped, single-stranded negative-sense RNA virus with an envelope G glycoprotein that is responsible for attachment, membrane fusion, and infection of host cells. ABLV is categorized in Group I of the NIAID Emerging and Re-emerging Diseases list, one circulating in flying foxes (fruit bats) and another in insectivorous bats. Each caused fatal rabies-like encephalitis in humans but with quite different incubation periods. Previously, we developed successful research programs on the henipaviruses (Hendra and Nipah) in our first support period, and many of the tools and expertise derived from those studies along with our collaborative ties with investigators in Australia now offers us the unique opportunity to initiate a new project within the MARCE-2 renewal. Here, we will address several areas related to ABLV binding, entry and infection of host cells, providing new data on this under-researched emerging virus as well as potentially uncovering new information that may be applicable to other viruses within the Mononegavirales, including the filoviruses (Ebola and Marburg) and the henipaviruses (Hendra and Nipah) - areas in which we are well experienced. We have long standing collaborations with leading investigators in Australia: Dr. Linfa Wang at the CSIRO and Dr. Ina Smith at the Public Health Virology unit of Queensland Health, Forensic and Scientific Services. The overall goal of our project is to understand the binding and infection process of ABLV and to identify viral and host cell targets for developing therapeutic intervention strategies. Specifically, we propose to: 1. Explore and characterize the host cell type and species tropisms of ABLV. 2. Develop and characterize soluble monomeric and trimeric forms of ABLV-G; and isolate and characterize human anti-G Fabs and domain antibody (dAb) using phage-display platforms. 3. We will conduct high-throughput RNAi screening for host cell factors required for ABLV infection and growth in cell culture.